Uncovering the Starr of the Cell - SUN and KASH Proteins: An Interview with Daniel A. Starr, PhD.

Dr. Daniel A. Starr is a cell biologist and professor in the department of Molecular and Cellular Biology at the University of California, Davis. He attained his B.S. in Biology and Ph.D. in Genetics from Colby College and Cornell University, respectively. As a Ph.D. student his research focused on the movement of chromosomes during mitosis using Drosophila melanogaster as his model organism. Currently, Dr. Starr focuses his research on nuclear positioning in eukaryotic cells using the model organism, Caenorhabditis elegans. His current project started in the early 2000s when he was a Postdoctoral fellow at the Han Lab in Boulder, Colorado, and is now continued in the Starr Lab at University of California, Davis. Dr. Starr’s efforts led to the discovery of the ANC-1 protein in the KASH proteins family. KASH proteins (Klarsicht, ANC-1, Syne homology) are membrane proteins residing in the outer membrane of the nuclear envelope. The KASH proteins are an essential component necessary for nuclear positioning. The SUN (Sad-1 and UNC-84) proteins are membrane proteins that reside in the inner nuclear membrane. Dr. Starr proposed a model in which the SUN and KASH proteins physically interact with one another to connect the two membranes via formation of a bridge that spans the nuclear envelope, essentially keeping the nucleus positioned (see Figure 1). This model was later on proven to be the correct model tested by many labs including Starr Lab. He is the recipient of the Basil O’Conner Scholar Award from March of Dimes and is a member of the American Society for Cell Biology. Dr. Daniel Starr is not only a well-reputed cell biologist, but also a committed mentor and professor of the UC Davis community. He is a favorite amongst his students and has been honored with the Faculty Development Award.

Daniel A. Starr, PhD

Professor of Molecular and Cellular Biology, University of California, Davis. The Starr Lab Website: https://starr.faculty.ucdavis.edu/

Figure 1

SUN-KASH protein interaction model. The SUN-KASH protein interaction bridges the inner and outer nuclear membrane. The adaptor domain on the cytoplasmic tail of KASH proteins varies depending on cell type and function.

Your efforts led to the discovery of the KASH proteins role in nuclear positioning. Were you in search of this particular interaction and can you elaborate more on the SUN-KASH domain relationship?

Using a forward-genetics approach I was unbiased in what I was searching for. All I knew was when either the SUN or KASH genes were mutated the nucleus was unable to move. I didn’t know what I was searching for but I knew there must be something localized to the nuclear surface – a membrane marker of a sort that was defining the outer nuclear membrane. Every membrane has a unique marker and perhaps this was the defining marker for the outer nuclear membrane. I proposed the model that SUN and KASH proteins interact with one another to bridge the nuclear envelope. I was the first to propose this idea, but soon after many labs started testing the model including my lab and it ended up being correct. Being a young scientist in the field I could not wrap my head around the thought of nothing being at the outer membrane to distinguish it from the other membranes present. Others in the field were skeptical of my hypothesis when I first started exploring the possibility, because it was a cell biology question no one had asked previously.

The SUN and KASH machinery is conserved and in my opinion important for the evolution of the first eukaryotes. The definition of a eukaryote is an organism with a nuclear envelope around its DNA (which becomes the nucleus). To have equal spacing between the outer and inner nuclear membrane, the SUN and KASH proteins are necessary, and there is a high probability this domain was produced in the last common eukaryotic ancestor and since then has been conserved. Different organisms use this SUN-KASH machinery differently, but one unanimous use is to distinguish the nucleus from the endoplasmic reticulum. Once the difference is marked various unique modules can be added to the cytoplasmic end of the KASH proteins. Some cells may want to interact with microtubule motors while others prefer intermediate filaments, myosins, or different kinesins. An appropriate adapter on the cytoplasmic end of the KASH proteins will follow for each interaction. The KASH proteins in plant cells have a different function, but they still mark the nucleus regardless of latter function.

Can you explain the importance of having nuclei positioned to a certain location in the cell? Why is the nucleus not a free floating organelle?

My lab uses C. elegans as our model organism and nuclei positioning may not be very important in the tissues we study in C. elegans, but it is critical in other eukaryotes. An obvious example of the importance of nuclei positioning is in skeletal muscle cells. Our skeletal muscle cells form when hundreds of single cells fuse together to form a long continuous myotube. The myotube runs the complete length of our muscle and the goal is to contract the entire myotube at once – having it function as one large cell. Even though the tube runs the entire length of the muscle, it has only one neuromuscular junction. The thin and thick filaments (myosin and actin respectively) are continuously sliding against each other during muscle contraction. To inhibit nuclei from interfering in this motion, the muscle cell localizes its nuclei to the periphery of this tube to not interfere with contraction. Energy investment for mRNA localization is an important factor the cell considers. If nuclei were unfavorably far from the epicenter of the cell, the amount of energy required to transport the mRNA long distances to where they are required will become a highly energy dependent process. The cells prefer to store that energy for other processes, and so the hundreds of nuclei in skeletal muscle are very evenly spread throughout the center of muscle cells.

Furthermore, some nuclei are able to (and therefore do) specialize which effects their positioning in the cell. To continue with the skeletal muscle cells, the nuclei that are directly under the neuromuscular junction make specialized mRNAs. An example of this is the Acetylcholine Receptor – a necessity for the cells directly under the neuromuscular junction, but not for the rest of the muscle cells. Post differentiation, nuclei are fixed in position to prevent movement into regions of the cell where the specialized nuclei are not needed. Most differentiated cells will polarize and localize their nuclei to a specific location (not all but most). The exact location of the nucleus is dependent on the cell type and function. Some cells may want the nucleus in a basal position while others prefer a more centralized position. The location is a part of the larger cell purpose/function. For example, the receptor cells that vibrate to sound in our ears have their nuclei at the base of the cell. In some hearing impaired patients, the position of the nuclei in receptor cells are found in a non-basal position.

Seeing how important the SUN-KASH domain is in nuclear positing, what does a mutation in this domain look like? Does the cell possess a correcting force if the nucleus is not in the correct location?

A null mutation in the SUN-KASH complex is embryonic lethal, therefore cannot be inherited from one generation to the next. SUN and KASH protein play a crucial role in the first mitosis event by pushing the male and female pronuclei towards each other. In the absence of this the embryo fails to develop further. Recently, weak mutations in SUN-KASH proteins have been linked to certain familial diseases that may be inherited. As of our knowledge today, these mutations may be a contributing factor in neurological diseases (schizophrenia and autism – minor contributing factors but important enough to mention), neuromuscular diseases, premature ageing diseases, and others still being discovered. Cancer cells have been found with a mutation in the SUN and/or KASH proteins and cells with a mutation in the SUN and/or KASH proteins in certain tissues have a 2 to 3 times higher probability of becoming cancerous cells. There is ongoing research with the mutants of these proteins and I imagine someone is working on developing a drug targeting this domain because it’s fascinating to see the amount of diseases that are being linked to the mutant forms of these proteins.

In terms of how nuclei are positioned we’ve established that mechanisms vary amongst cell types. In mature skeletal muscle cells, nuclei are out in the periphery, and if they are in the interior center then they are young cells that will eventually migrate the nuclei out to the periphery. In a stressful environment or after the cell has undergone damage (ex: weightlifting), nuclei migrate back to the center momentarily and back out once recovered. Nuclei permanently anchored in the center of the myotube are a hallmark of disease or muscle damage. Healthy cells can recover from the damage, migrating the nuclei back into the periphery over the course of a few days.

We know at this point that nuclear location varies amongst cells, but we have yet to discover all the mechanisms that guide nuclear positioning. This process is not as conserved and universal as the SUN-KASH machinery present once the nuclei are in the correct location. A layer of complexity is added because not all nuclear movements are done via SUN-KASH proteins. Some cells use cytoskeletal forces and other ways depending on what system is being observed. An additional mechanism identified is an actin-myosin net behind the nucleus that contracts and pushes the nucleus forward independent of SUN and KASH proteins. There is a lot to be discovered and as we dig deeper, more questions rise.

Now that you have answered your initial research question, what is next for Starr Lab?

A part of being a cell biologist is the question is never really answered because you keep uncovering new layers and keep going further into details. A recent paper that we published dove into the minute details of SUN-KASH proteins characterizing, which amino acids interact with which, and how mutations in those amino acids impact the cell [1]. The SUN and KASH proteins relationship was found about 15 years ago now, but we are still unveiling details about the model every day. I’m now interested in what the cytoplasmic and nucleuplasmic domains of SUN-KASH complexes interact with and how such interactions play roles in cellular processes.

Another question that has been crossing my mind recently is, how is the nucleus flexible enough to squeeze through narrow “holes”? Certain sets of nuclei have to migrate through small spaces via contraction and squeezing through. We know at this point the nucleus is the largest and stiffest organelle in the cell. The efficiency and ability of cells to pass through the narrow opening is therefore dependent on how well nuclei pass through. The SUN and KASH proteins are involved in the process of nuclear movement. We have little pieces of the puzzle that need to be placed correctly to understand the mechanism exactly. Recently a second, actin based, genetic pathway was identified that works in conjunction with SUN and KASH proteins, but its complete function is still undetermined. We think nuclear squeezing through constrictions is important in cancer because metastasizing cells migrate through the extracellular matrix and need flexibility to do so. In C. elegans, cell migration is a normal developmental step, and C. elegans use both the SUN-KASH domain and the secondary pathway. I would like to study in more depth this secondary pathway and connect the dots to explain the role it plays in cell migration. Recently my lab has also been going into a different territory of how mechanical forces are transmitted from the rest of the cell to the nucleus. One of my graduate students is interested in seeing how these forces effect gene transcription and regulation inside the nucleus.

To answer what our long-term goal is, I would say it’s to understand how organelles interact with each other and how they interact with the whole cell to set up the global cellular architecture. One student is looking at other organelles in the cell of these mutants and has observed many mislocalized organelles, indicating the nuclear envelope might regulate the whole, global cell architecture.